Expression and transport characterisation of the wheat low-affinity cation transporter (LCT1) in the methylotrophic yeast Pichia pastoris

Biochemical and Biophysical Research Communications
Eugene DiatloffStephen K Roberts

Abstract

The low-affinity cation transporter (LCT1) from wheat (Triticum aestivum) was expressed in the methylotrophic yeast Pichia pastoris and its transport characteristics studied employing Ca(45) and Cd(109). A clone (LCT1#3) with the highest uptake of 14pmol of Ca/10(6)cells/10min when exposed to 100microM Ca(45) was chosen for further Ca(45) and Cd(109) transport characteristics. We report for the first time a K(m) for Ca by LCT1 of 0.43+/-0.15mM Ca activity which confirms LCT1 to be a low affinity transporter. Interestingly, the expression of LCT1 in Pichia resulted in reduced Cd(109) uptake compared to wild type cells, when cells were exposed to >or=60microM Cd. This is the first report of the ability of a heterologously expressed transporter to reduce the activity of endogenous transporter proteins to transport Cd. To our knowledge, this is the first demonstration of functional expression of a plant ion transporter using P. pastoris.

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Citations

Aug 9, 2012·The Journal of Membrane Biology·Ulrich TerpitzVladimir L Sukhorukov
Jun 18, 2009·Plant Cell Reports·Chang Quan Wang, Heng Song
Aug 13, 2009·International Journal of Plant Genomics·Filiz Yesilirmak, Zehra Sayers
May 13, 2009·The International Journal of Biochemistry & Cell Biology·Jose R Peralta-VideaJorge Gardea-Torresdey
Apr 23, 2010·Annals of Botany·Simon Conn, Matthew Gilliham

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